T-lymphocytes can be activated via the CD2 and CD3 receptors through the use of specific M.Ab and mitogenic lectins. A host of second messengers are required to set the stage for subsequent autocrine proliferation through secretion of IL-2 and expression of its receptor. The long term objective is an understanding of the importance of phospholipid metabolism, C-kinase activity and cell CA+2 along these activation pathways with the view to define target areas for response modification in disease. An understanding of these mechanisms may also explain the pathophysiological basic of certain immuno-deficiencies. The specific aims are therefore to use M.Ab and mitogenic lectins to define the cell biological role of C-kinase and CA+2 i.t.o. I1-2 receptor expression, substrate phos-phorylation, receptor modulation and expression of mRNA for c-fos, c-myc, IL-2, gamma-IFN, and the IL-2 receptor. These studies will also look at some of the processes which precedes the activation of C-kinase, namely PI turnover, IP3, release, CA+2 flux and receptor aggregation. This may reveal differences in the potency by which second messengers are generated, for instance between lectins and anti-T3 M.Ab. The influence of chemical modulators/drugs on these processes will also be examined. The signal transducing role of receptor related G-binding proteins will be investigated in terms of their potential for stimulating both adenylate cyclase and phospholipase C activities. The importance of in vivo autophosphorylation, translocation and proteolytic activation of C-kinase will be addressed. Finally, the important interaction of C-kinase with the tyrosine kinase, pp56 Tck, will be studied with a view to understanding the possible link between membrane phospholipid metabolism and mitogenic events. In order to perform these studies, the following methodology has been developed: protein phosphorylation studies, immunoprecipitation of specific phosphorylated substrates, SDS- PAGE and autoradiography, phospho-amino acid analysis, TLC of phospholipid extracts, ion exchange chromatography to measure IP3 release and cytoplasmic dot-blot analysis for mRNA. Intact cell responses are monitored by flow cytometry, 3H-Td uptake, IL-2 secretion and Quin 2 fluorescence.